1. Field of the Invention
The present invention relates to cell-associated glucosyltransferase which reacts with sucrose and catalyzes a reaction of synthesizing water-insoluble glucans, to a method of purifying this enzyme and to a method of producing this enzyme.
The present invention also relates to an antibody having immunological activity against Streptococcus mutans which is known as pathogenic bacteria inducing dental-caries and to a dental caries prophylactic composition which contains said antibody as an effective component.
2. Description of the Prior Art
In view of dental caries prophylaxis, a number of investigations have been undertaken to elucidate the properties of S. mutans known as causative bacteria for dental caries and the mechanism of involvement of said bacteria in the development of dental caries.
As to S. mutans strains which are involved in the development of dental caries, eight strains, being classified serologically into serotypes a to h, have now been found. Of these serotypes, two subgroups have been defined on the basis of the serological analogy, serotypes d and g in one group and serotypes c, e and f in the other group.
It is already known that the important process in the mechanism of the involvement of S. mutans in developing dental caries is the process of producing adherent, water-insoluble glucans from sucrose, in which glucosyltransferase (hereinafter referred to as GTF) produced by S. mutans plays a part.
This GTF produced by S. mutans can be found either in the culture supernatant or on the cell surface in an associated form.
For example, in the culture of strains of serotype d or g, serologically analogous one another, in the absence of sucrose, GTF is present mostly in the culture supernatant, from which three kinds of GTF, namely GTF-S1, GTF-S2 and GTF-I, have now been isolated and purified (S stands for a water-soluble glucan synthesizing enzyme and I stands for a water-insoluble glucan synthesizing enzyme). These three kinds of GTF cooperatively produce adherent, water-insoluble glucans (see, for example, "Synthesis of glucans by Streptococcus mutans and adherence mechanisms", T. Koga, Jap. J. Bacteriol., 41 (4), 679, 1986).
Further, in the culture of the strains of serotypes d and g in the presence of sucrose, it is known that the aforementioned three kinds of GTF are mostly bound to the cells by binding to glucans to give cell-associated forms (Hamada, S. and Slade, H. D., Archs Oral. Biol., 24, 399-402, 1979).
On the other hand, GTF-S and GTF-I have been isolated also from the culture supernatant of the strains of serotypes c, e and f, serologically analogous one another (Kuramitsu, H. K. and Wondrack, L., Infect. Immun., 42, 763-770, 1983).
However, there has been no single report that water-insoluble, adherent glucans can be synthesized in joint action of these GTF-S and GTF-I.
Furthermore, attempts have been made to develop methods or agents for preventing dental caries, making S. mutans a target organism. A known example of the method of preventing dental caries by controlling the colonization of S. mutans in the oral cavity (especially by controlling the adherence to tooth surfaces) is the method for dental caries prevention by using antibodies which have immunological activity against S. mutans. Namely, a method of preventing the colonization of S. mutans in the oral cavity by using milk obtained from cows which are immunized by cells of S. mutans has been disclosed in British Patent Specification No. 1505513, and a dental caries preventive agent which is a combination of antiserum and/or milk obtained from the mammalian having been immunized by a fraction such as cell walls recovered from the culture of S. mutans and one or more of the synergists selected from the group consisting of glucosyltransferase inhibitors, proteases and dextranases has been disclosed in the specification of Japanese Patent Laid Open No. 60-38327.
However, the effect of preventing dental caries by the antibodies having immunological activity against S. mutans is not always satisfactory. Furthermore, since the antibodies are conventionally prepared from milk or antiserum of immunized mammalians, in many cases, they are not practically produced owing to such disadvantages as inappropriateness to mass production or high production cost.